Bibliographic details of the publications referred to by author in this specification are collected at the end of the description.
Reference to any prior art in this specification is not, and should not be taken as, an acknowledgment or any form of suggestion that this prior art forms part of the common general knowledge in any country.
Psoriasis and other similar conditions are common and often distressing proliferative and/or inflammatory skin disorders affecting or having the potential to affect a significant proportion of the population. The condition arises from over proliferation of basal keratinocytes in the epidermal layer of the skin associated with inflammation in the underlying dermis. Whilst a range of treatments have been developed, none is completely effective and free of adverse side effects. Although the underlying cause of psoriasis remains elusive, there is some consensus of opinion that the condition arises at least in part from over expression of local growth factors and their interaction with their receptors supporting keratinocyte proliferation via keratinocyte receptors which appear to be more abundant during psoriasis.
One important group of growth factors are the dermally-derived insulin-like growth factors (IGFs) which support keratinocyte proliferation. In particular, IGF-I and IGF-2 are ubiquitous polypeptides each with potent mitogenic effects on a broad range of cells. Molecules of the IGF type are also known as “progression factors” promoting “competent” cells through DNA synthesis. The IGFs act through a common receptor known as the Type I receptor or IGF-IR, which is tyrosine kinase linked. They are synthesized in mesenchymal tissues, including the dermis, and act on adjacent cells of mesodermal, endodermal or ectodermal origin. The regulation of their synthesis involves growth hormone (GH) in the liver, but is poorly defined in most tissues (Sara, Physiological Reviews 70: 591-614, 1990).
Particular proteins, referred to as IGF binding proteins (IGFBPs), appear to be involved in autocrine/paracrine regulation of tissue IGF availability (Rechler and Brown, Growth Regulation 2: 55-68, 1992). Six IGFBPs have so far been identified. The exact effects of the IGFBPs is not clear and observed effects in vitro have been inhibitory or stimulatory depending on the experimental method employed (Clemmons, Growth Regn. 2:80, 1992). There is some evidence, however, that certain IGFBPs are involved in targeting IGF-I to its cell surface receptor.
Skin, comprising epidermis and underlying dermis, has GH receptors on dermal fibroblasts (Oakes et al., J. Clin. Endocrinol. Metab. 73: 1368-1373, 1992). Fibroblasts synthesize IGF-1 as well as IGFBPs-3, -4, -5 and -6 (Camacho-Hubner et al., J. Biol. Chem. 267: 11949-11956, 1992) which may be involved in targeting IGF-1 to adjacent cells as well as to the overlaying epidermis. The major epidermal cell type, the keratinocyte, does not synthesize IGF-I, but possesses IGF-I receptors and is responsive to IGF-I (Neely et al., J. Inv. Derm. 96: 104, 1991).
In the last decade, there have been many reports of the use of antisense oligonucleotides to explore gene function and in the development of nucleic acid based drugs. Antisense oligonucleotides inhibit mRNA translation via a number of alternative ways including destruction of the target mRNA through RNaseH recruitment, or interference with RNA processing, nuclear export, folding or ribosome scanning. More recently, a better understanding of intracellular sites of action of the various antisense modalities and improvements in oligonucleotide chemistry have increased the number of reports of validated expression inhibition.
In work leading up to the present invention, the inventors focused on the use of the antisense approach to inhibit the growth of human epidermal keratinocytes, particularly in human epidermal growth disorders such as psoriasis. Psoriasis is a common and disfiguring skin condition associated with severe epidermal hyperplasia. Existing psoriasis therapies are only partially effective, however, treatments targeting the epidermis have shown promise (Jensen et al., Br. J. Dermatol. 139: 984-991, 1998; van de Kerkhof, Skin Pharmacol. Appl. Skin Physiol. 11: 2-10, 1998). One strategy is to develop antisense inhibitors of IGF-IR expression and to use these to block IGF-I-stimulated cell division and survival in the epidermis.
The IGF-IR is a tyrosine kinase linked cell surface receptor (Ullrich et al., EMBO J. 5: 2503-2512, 1986) that regulates cell division, transformation and apoptosis in many cell types (LeRoith et al., Endocr. Rev. 16: 143-163, 1995; Rubin and Baserga, Laboratory Investigation 73: 311-331, 1995). Human epidermal keratinocytes are highly responsive to IGF-IR activation (Ristow and Messmer, J. Cell Physiol. 137: 277-284, 1988; Neely et al., J. Invest. Dermatol. 96: 104-110, 1991; Wraight et al., J. Invest. Dermatol. 103: 627-631, 1994) and several studies point to an important role for IGF-1R activation in the pathogenesis of psoriasis (Krane et al., J. Invest. Dermatol. 96: 419-424, 1991; Krane et al., J. Exp. Med. 175: 1081-1090, 1992; Ristow, Growth Regul. 3: 129-137, 1993; Hodak et al., J. Invest. Dermatol. 106: 564-570, 1996; Xu et al., J. Invest. Dermatol. 106: 109-112, 1996; Ristow, Dermatology 195: 213-219, 1997; Wraight et al., J. Invest. Dermatol. 108: 452-456, 1997). The IGF-IR has been targeted previously by antisense approaches in fibroblasts, haemopoietic cells and glioblastoma cells to investigate its role in transformation and cell cycle progression (Pietrzkowski et al., Mol. Cell. Biol. 12: 3883-3889, 1992; Porcu et al., Mol. Cell. Biol. 12: 5069-5077, 1992; Reiss et al., Oncogene 7: 2243-2248, 1992; Resnicoff et al., Cancer Res. 54: 2218-2222, 1994).
The identification of propynylated phosphorothioate oligonucleotides have been reported which are capable of reducing IGF-IR mRNA levels when efficiently delivered to the keratinocyte nucleus (White et al., Antisense Nucleic Acid Drug Dev. 10: 195-203, 2000; Wraight et al., Nat. Biotechnol. 18: 521-526, 2000). These oligonucleotides were also effective at reducing IGF-I binding, receptor activation and cell proliferation in vitro and epidermal proliferation in vivo (Wraight et al., 2000, supra).
Propyne-modified phosphorothioate oligonucleotides were selected (Flanagan et al., Nat. Biotechnol. 14: 1139-1145, 1996b; Flanagan and Wagner, Mol. Cell. Biochem. 172: 213-225, 1997) because their increased affinity for target mRNA allows mRNA inhibition with lower concentrations (Wagner et al., 1993, supra) and shorter oligonucleotide length (Flanagan et al., Nucleic Acids Res. 24: 2936-2941, 1996a) than unmodified phosphorothioates, theoretically reducing the incidence of aptameric effects on target cells.
Whilst success has been demonstrated with the propyne-modified phosphorothioate oligonucleotides, alternative chemistries need to be considered to reduce toxicity, increase stability, increase specificity profile, improve penetration and/or to enhance potency and biological, chemical or physical properties. Oligonucleotides of alternative chemistries can also provide other advantages including known large scale manufacture, human clinical development knowhow, and/or known approval as drugs.